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S Wissing, T Laube, C Theiss, KP Steuhl, D Meller; Reduced Keratocyte Loss after Laser Epithelial Keratomileusis (LASEK) in Comparison to Photorefractive Keratectomy (PRK) in Rabbits . Invest. Ophthalmol. Vis. Sci. 2002;43(13):4242.
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Purpose: To compare corneal wound healing with special regard to keratocyte loss in corneal stroma after LASEK and conventional PRK in rabbits. Methods: LASEK and PRK were performed in rabbits and studied after 1, 3, 10 and 21 days (each group n=3-5). In all eyes, hemidesmosomes of corneal epithelium were weakened with 30% ethanol for 3.5 min. In the PRK group, the whole epithelium was removed, but in the LASEK group epithelial flap was repositioned and left to adhere to the underlying stromal bed. Excimer photoablation was performed unilaterally with a 6.0mm ablation zone and 80µm depth equivalent to -6.0 dpt using the SUMMIT APEX PLUS laser with photorefractive keratectomy (PRK) mode. Keratocyte loss was analyzed using a DNA fragmentation detecting TUNEL assay. TUNEL positive keratocytes were counted in 4 different high power fields. Monoclonal antibodies to single stranded DNA (ssDNA) which has been observed recently in cells undergoing apoptosis and to α-smooth muscle actin which has been found in migrating keratocytes were applied for further analysis. Results: One day after LASEK, eyes showed minor epithelial defects, whereas PRK treated eyes developed a distinct wound. In general, corneal wound healing occured faster after LASEK than PRK. After 10 days a smooth, stable and optically transparent cornea was documented in all eyes. Numerous TUNEL positive keratocytes predominantely located in the anterior corneal stroma were noted 1 day after PRK which diminished slightly after 3 days. LASEK treated eyes revealed significant less TUNEL positive keratocytes at these time points and showed a delayed transient increase of TUNEL positive cells 3 days after laser photoablation. At 10 days, the number of TUNEL positive keratocytes decreased in both surgical groups but remained significantly higher after PRK than LASEK. The ssDNA antibody labeled a more dense accumulation of positive cells presumably representing apoptotic cells in PRK than LASEK. In both surgical methods, α-smooth muscle actin was not found at any time point analyzed. Conclusion: LASEK does induce significantly less keratocyte loss than PRK and supports wound healing in the acute phase after laser photoablation. Further studies such as electron microscopy are currently carried out to investigate if keratocyte loss is mediated by apoptosis. LASEK might offer the possibility to treat higher myopia with reduced risk of developing complications as known to occur in PRK.
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